Hyaluronic acid (HA) is a member of a class of polymers known as glycosaminoglycans. HA is a long chain linear polysaccharide and is usually present as a sodium salt having the molecular formula (C14H20NNa11)n where n may vary according to the source of the HA and the method of isolating the HA. Molecular weights of HA of up to 14×106 have been reported.
HA and its salts may be isolated from many sources including the human umbilical cord, rooster combs and nearly all connective matrices of vertebrate organisms. HA is also a capsular component of bacteria such as streptococci and may therefore be obtained by fermentation methods such as reported in U.S. Pat. No. 5,411,874 (Fermentech Ltd).
HA is non-immunogenic and therefore has great potential in medicine. Because of its visco-elastic properties, HA having a high molecular weight (over 1 million) has been found to be particularly useful in a variety of clinical fields, including wound treatment, ophthalmic surgery, orthopedic surgery and drug delivery. HA is also potentially useful in a variety of non-medical fields, including cosmetic applications.
However, one drawback to administering HA to humans is that HA is degraded by enzymes such as hyaluronidase and free radicals found in the human body. Furthermore, HA is soluble in water at room temperature, which may also make it less suited to certain applications.
Various attempts have been made to prepare more stable forms of HA, in particular, by cross-linking the HA molecules. For example, hydroxyl groups have been cross-linked via an ether linkage and carboxyl groups via an ester linkage. HA has been cross-linked at pH levels less than 9 at which ester bonds form via carboxyl groups, and at pH levels greater than 9 at which ether bonds form via hydroxyl groups. The present inventors have found that ether bonds may be beneficial because these bonds are more resistant to physiological degradation.
A number of documents report a variety of methods of cross-linking HA gels. For example, U.S. Pat. No. 4,582,865 (Biomatrix Inc) reports cross-linked gels of HA formed by cross-linking HA (either by itself or mixed with other hydrophilic polymers) using divinyl sulfone as the cross-linking agent.
U.S. Pat. No. 5,827,937 (Agerup) reports polysaccharide gel compositions prepared by forming an aqueous solution of the polysaccharide, initiating cross-linking in the presence of a polyfunctional cross-linking agent, sterically hindering the cross-linking reaction from being terminated before gelation occurs (e.g. by diluting the solution) and then reintroducing sterically unhindered conditions (e.g. by evaporating the solution) so as to continue the cross-linking such that a viscoelastic gel is formed. The cross-linking in this method may be performed under alkaline or acidic conditions.
WO 00/46253 (Fermentech Ltd) reports cross-linking HA with other polymers by two different types of cross-linking bonds. The formation of different types of bonds is achieved by cross-linking via different functional groups. For example, one type of bond may be formed by cross-linking via hydroxyl groups, and a different functional bond may be formed by cross-linking via carboxyl groups.
WO 87/07898 reports reacting a polysaccharide with a polyfunctional epoxide, removing excess epoxide and employing a drying operation to cross-link the polysaccharide into a film, powdered material or similar dry product.
U.S. Pat. No. 4,963,666 (Pharmacia) reports a process in which a polysaccharide is monosubstituted with a cross-linking agent at low concentration under alkaline conditions to form ether linkages. The mixture is washed to pH 5.5 inducing some ester linkages and then, in one example, concentrated by slow evaporation to complete cross-linking with ester linkages. In another example, the pH is increased by the addition of ammonia, and then slowly evaporated to complete the cross-linking with primarily ether linkages and some ester linkages.
Although attempts have been made to improve the properties of cross-linked HA, it would be beneficial to provide cross-linked HA gels having improved degradation characteristics when administered to a patient.